Transformation of algae for increasing lipid production

Abstract

The present disclosure provides novel proteins that when over expressed in algae result in an increase or change in fatty acid and/or glycerol lipid production and/or accumulation, without a substantial decrease in the growth rate of the alga or the break down of algal components, such as chlorophyll. The present disclosure also describes methods of using the novel proteins to increase or change the production and/or accumulation of fatty acids and/or glycerol lipids in algae. In addition, these proteins are useful tools in obtaining information about the fatty acid and triacyglyceride (TAG) synthetic pathways in algae.

What is claimed is: 
     
       1. A non-vascular photosynthetic organism transformed with a polynucleotide, wherein the polynucleotide comprises: (a) a nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4; or (b) a nucleic acid sequence with at least 95%, at least 98%, or at least 99% sequence identity to the nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4, wherein the transformed organism has an increased lipid content than the untransformed organism. 
     
     
       2. The transformed organism of  claim 1 , wherein the nucleic acid sequence encodes a polypeptide comprising an amino add sequence of SEQ ID NO: 52 or SEQ ID NO: 51, or at least 95%, at least 98%, or at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51. 
     
     
       3. The transformed organism of  claim 1 , wherein the transformed organism is an alga or a photosynthetic bacterium. 
     
     
       4. The transformed organism of  claim 3 , wherein the photosynthetic bacterium is a cyanobacterium. 
     
     
       5. The transformed organism of  claim 3 , wherein the alga is a microalga. 
     
     
       6. The transformed organism of  claim 1 , wherein the increase is shown by a change in the amount of total lipids. 
     
     
       7. The transformed organism of  claim 1 , wherein the increase is shown by a change in the amount of total gravimetric lipids. 
     
     
       8. The transformed organism of  claim 1 , wherein the increase is shown by a change in percent lipids by use of hexane extraction. 
     
     
       9. The transformed organism of  claim 1 , wherein the increase is shown by a change in percent lipids by use of methyl tert-butyl ether (MTBE) extraction. 
     
     
       10. The transformed organism of  claim 1 , wherein the lipid is a triacylglycerol (TAG), a diacylglycerol (DAG), a glycosylglycerol, a neutral storage lipid, a polar lipid, a fat, a wax, a sterol, a fat-soluble vitamin, a monoglyceride, a diglyceride, a triglyceride, a phospholipid, a fatty acyl, a glycerolipid, a glycerophospholipid, a sphingolipid, a polyketide, a sterol lipid, a prenol lipid, a fatty acid, a fatty acid derivative, a sterol-containing metabolite, or a combination of any two or more thereof. 
     
     
       11. The transformed organism of  claim 1 , wherein the increase is determined by use of methyl-tert-butyl ether (MTBE) extraction. 
     
     
       12. The transformed organism of  claim 1 , wherein the increase is determined by use of a lipid dye. 
     
     
       13. The transformed organism of  claim 12 , wherein the lipid dye is Bodipy, LipidTOXgreen, or Nile Red. 
     
     
       14. The transformed organism of  claim 1 , wherein the lipid is stored in a lipid body, a cell membrane, an inter-thylakoid space, or a plastoglubuli of the transformed organism. 
     
     
       15. A method of increasing lipid content of a non-vascular photosynthetic organism, comprising: i) transforming the organism with a polynucleotide comprising a nucleotide sequence encoding a protein expressed in the transformed organism, wherein expression of the protein results in an increase in the lipid content of the transformed organism as compared to the untransformed organism, and wherein the nucleotide sequence comprises, (a) a nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4, or (b) a nucleic acid sequence with at least 95%, at least 98%, or at least 99% sequence identity to the nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4. 
     
     
       16. The method of  claim 15 , wherein the protein comprises an amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51, or encodes a polypeptide comprising at least 95%, at least 98%, or at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51. 
     
     
       17. The method of  claim 15 , further comprising determining the change increase by use of methyl-tert-butyl ether (MTBE) extraction or use of a lipid dye. 
     
     
       18. The transformed organism of  claim 1 , wherein said polynucleotide comprises: (a) a nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4; or (b) a nucleic acid sequence with at least 95% 98% sequence identity to the nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4. 
     
     
       19. The transformed organism of  claim 2 , wherein said nucleic acid sequence or nucleotide sequence encodes a polypeptide comprising an amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51, or at least 98% sequence identity to the amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51. 
     
     
       20. The method of  claim 15 , wherein nucleotide sequence comprises, (a) a nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4, or (b) a nucleic acid sequence with at least 98% sequence identity to the nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4. 
     
     
       21. The method of  claim 16 , wherein said comprising an amino acid sequence of SEQ ID NO: 52 or encodes a polypeptide comprising at least 98% sequence identity to the amino acid sequence of SEQ ID NO: 52. 
     
     
       22. The transformed organism of  claim 1 , wherein said polynucleotide comprises: (a) a nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4; or (b) a nucleotide sequence with at least 99% sequence identity to the nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4. 
     
     
       23. The transformed organism of  claim 2 , wherein said nucleic acid sequence or nucleotide sequence encodes a polypeptide comprising an amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51, or at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 52 or SEQ ID NO: 51. 
     
     
       24. The method of  claim 15 , wherein nucleotide sequence comprises, (a) a nucleic acid sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4, or (h) a nucleic acid sequence with at least 99% sequence identify to the nucleic add sequence of SEQ ID NO: 9, SEQ ID NO: 12, or SEQ ID NO: 4. 
     
     
       25. The method of  claim 16 , wherein said polypeptide comprising an amino acid sequence of SEQ ID NO: 52 or at least 98% sequence identity to the amino acid sequence of SEQ ID NO: 52.